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1.  Castleman’s disease- a diagnostic dilemma 
Castleman’s disease is a benign lymphoproliferative disease characterised by hyperplasia of lymphoid follicles. It can affect any lymph nodes in the body. Here we describe a caucasian patient who presented with six months history of shortness of breath with CT scan confirming an 8 cm segment of consolidated lung in left hilum. PET scan revealed a mass measuring 68x80x55 mm extending from the left hilum out into the left upper lobe containing area of calcification with SUV max 4.8. The differential diagnosis included atypical sequestration, hamartoma and primary lung malignancy. The patient underwent left video assisted enucleation of the lesion. The histology confirmed the diagnosis of Castleman’s disease.
doi:10.1186/s13019-014-0170-0
PMCID: PMC4255971  PMID: 25431102
Benign lung lesion; Castleman’s disease and VATS
2.  EPIQ Review 
Paediatrics & Child Health  2014;19(4):176.
BACKGROUND:
A checklist that promotes compliance with aseptic technique during line insertion is a component of many care bundles aimed at reducing nosocomial infections among intensive care unit patients.
OBJECTIVE:
To determine whether the use of bundled interventions that include a checklist during central-line insertions reduces catheter-related bloodstream infections in intensive care unit patients.
METHODS:
A literature review was performed using methodology adapted from the American Heart Association’s International Liaison Committee on Resuscitation.
RESULTS:
Seventeen cohort studies were included. Thirteen studies were supportive of the intervention, while four were neutral. Infection rates ranged from 1.6 to 10.8 per 1000 central-line days in control groups, and from 0.0 to 3.8 per 1000 central-line days in the intervention groups.
CONCLUSION:
There is fair evidence to recommend the use of care bundles that include a checklist during central-line insertion in intensive care unit patients to reduce the incidence of catheter-related bloodstream infections.
PMCID: PMC4028639  PMID: 24855410
Catheters; Catheter-related infections; Checklist; Health care; Indwelling/adverse events; Infection control/methods; Intensive care units; Quality assurance
3.  Use of bundled interventions, including a checklist to promote compliance with aseptic technique, to reduce catheter-related bloodstream infections in the intensive care unit 
Paediatrics & Child Health  2014;19(4):e20-e23.
BACKGROUND:
A checklist that promotes compliance with aseptic technique during line insertion is a component of many care bundles aimed at reducing nosocomial infections among intensive care unit patients.
OBJECTIVE:
To determine whether the use of bundled interventions that include a checklist during central-line insertions reduces catheter-related bloodstream infections in intensive care unit patients.
METHODS:
A literature review was performed using methodology adapted from the American Heart Association’s International Liaison Committee on Resuscitation.
RESULTS:
Seventeen cohort studies were included. Thirteen studies were supportive of the intervention, while four were neutral. Infection rates ranged from 1.6 to 10.8 per 1000 central-line days in control groups, and from 0.0 to 3.8 per 1000 central-line days in the intervention groups.
CONCLUSION:
There is fair evidence to recommend the use of care bundles that include a checklist during central-line insertion in intensive care unit patients to reduce the incidence of catheter-related bloodstream infections.
PMCID: PMC4028651  PMID: 24855420
Catheters; Catheter-related infections; Checklist; Health care; Indwelling/adverse events; Infection control/methods; Intensive care units; Quality assurance
4.  Inhibition of a TREK-like K+ channel current by noradrenaline requires both β1- and β2-adrenoceptors in rat atrial myocytes 
Cardiovascular Research  2014;104(1):206-215.
Aims
Noradrenaline plays an important role in the modulation of atrial electrophysiology. However, the identity of the modulated channels, their mechanisms of modulation, and their role in the action potential remain unclear. This study aimed to investigate the noradrenergic modulation of an atrial steady-state outward current (IKss).
Methods and results
Rat atrial myocyte whole-cell currents were recorded at 36°C. Noradrenaline potently inhibited IKss (IC50 = 0.90 nM, 42.1 ± 4.3% at 1 µM, n = 7) and potentiated the L-type Ca2+ current (ICaL, EC50 = 136 nM, 205 ± 40% at 1 µM, n = 6). Noradrenaline-sensitive IKss was weakly voltage-dependent, time-independent, and potentiated by the arachidonic acid analogue, 5,8,11,14-eicosatetraynoic acid (EYTA; 10 µM), or by osmotically induced membrane stretch. Noise analysis revealed a unitary conductance of 8.4 ± 0.42 pS (n = 8). The biophysical/pharmacological properties of IKss indicate a TREK-like K+ channel. The effect of noradrenaline on IKss was abolished by combined β1-/β2-adrenoceptor antagonism (1 µM propranolol or 10 µM β1-selective atenolol and 100 nM β2-selective ICI-118,551 in combination), but not by β1- or β2-antagonist alone. The action of noradrenaline could be mimicked by β2-agonists (zinterol and fenoterol) in the presence of β1-antagonist. The action of noradrenaline on IKss, but not on ICaL, was abolished by pertussis toxin (PTX) treatment. The action of noradrenaline on ICaL was mediated by β1-adrenoceptors via a PTX-insensitive pathway. Noradrenaline prolonged APD30 by 52 ± 19% (n = 5; P < 0.05), and this effect was abolished by combined β1-/β2-antagonism, but not by atenolol alone.
Conclusion
Noradrenaline inhibits a rat atrial TREK-like K+ channel current via a PTX-sensitive mechanism involving co-operativity of β1-/β2-adrenoceptors that contributes to atrial APD prolongation.
doi:10.1093/cvr/cvu192
PMCID: PMC4174890  PMID: 25205295
Background K+ current; Beta-adrenoceptor; K2P channel; Steady-state outward current; Osmotic stretch; TREK-1; Arachidonic acid
5.  Insulin Protects Pancreatic Acinar Cells from Palmitoleic Acid-induced Cellular Injury* 
The Journal of Biological Chemistry  2014;289(34):23582-23595.
Background: Palmitoleic acid is a major pancreatitis-inducing agent.
Results: Insulin protected cells from palmitoleic acid (POA)-induced ATP depletion, inhibition of the plasma membrane calcium pump (PMCA), cytotoxic Ca2+ overload and necrosis.
Conclusion: Insulin protects against acinar cell injury induced by pancreatitis-inducing agents.
Significance: This provides an important therapeutic strategy for treating pancreatitis with insulin therapy.
Acute pancreatitis is a serious and sometimes fatal inflammatory disease where the pancreas digests itself. The non-oxidative ethanol metabolites palmitoleic acid (POA) and POA-ethylester (POAEE) are reported to induce pancreatitis caused by impaired mitochondrial metabolism, cytosolic Ca2+ ([Ca2+]i) overload and necrosis of pancreatic acinar cells. Metabolism and [Ca2+]i are linked critically by the ATP-driven plasma membrane Ca2+-ATPase (PMCA) important for maintaining low resting [Ca2+]i. The aim of the current study was to test the protective effects of insulin on cellular injury induced by the pancreatitis-inducing agents, ethanol, POA, and POAEE. Rat pancreatic acinar cells were isolated by collagenase digestion and [Ca2+]i was measured by fura-2 imaging. An in situ [Ca2+]i clearance assay was used to assess PMCA activity. Magnesium green (MgGreen) and a luciferase-based ATP kit were used to assess cellular ATP depletion. Ethanol (100 mm) and POAEE (100 μm) induced a small but irreversible Ca2+ overload response but had no significant effect on PMCA activity. POA (50–100 μm) induced a robust Ca2+ overload, ATP depletion, inhibited PMCA activity, and consequently induced necrosis. Insulin pretreatment (100 nm for 30 min) prevented the POA-induced Ca2+ overload, ATP depletion, inhibition of the PMCA, and necrosis. Moreover, the insulin-mediated protection of the POA-induced Ca2+ overload was partially prevented by the phosphoinositide-3-kinase (PI3K) inhibitor, LY294002. These data provide the first evidence that insulin directly protects pancreatic acinar cell injury induced by bona fide pancreatitis-inducing agents, such as POA. This may have important therapeutic implications for the treatment of pancreatitis.
doi:10.1074/jbc.M114.589440
PMCID: PMC4156068  PMID: 24993827
Calcium; Calcium ATPase; Calcium Transport; Insulin; Pancreas; PMCA; Pancreatitis
6.  Joint iris boundary detection and fit: a real-time method for accurate pupil tracking 
Biomedical Optics Express  2014;5(8):2458-2470.
A range of applications in visual science rely on accurate tracking of the human pupil’s movement and contraction in response to light. While the literature for independent contour detection and fitting of the iris-pupil boundary is vast, a joint approach, in which it is assumed that the pupil has a given geometric shape has been largely overlooked. We present here a global method for simultaneously finding and fitting of an elliptic or circular contour against a dark interior, which produces consistently accurate results even under non-ideal recording conditions, such as reflections near and over the boundary, droopy eye lids, or the sudden formation of tears. The specific form of the proposed optimization problem allows us to write down closed analytic formulae for the gradient and the Hessian of the objective function. Moreover, both the objective function and its derivatives can be cast into vectorized form, making the proposed algorithm significantly faster than its closest relative in the literature. We compare methods in multiple ways, both analytically and numerically, using real iris images as well as idealizations of the iris for which the ground truth boundary is precisely known. The method proposed here is illustrated under challenging recording conditions and it is shown to be robust.
doi:10.1364/BOE.5.002458
PMCID: PMC4132980  PMID: 25136477
(100.0100) Image processing; (100.4999) Pattern recognition, target tracking; (150.0150) Machine vision; (150.1135) Algorithms; (170.4470) Ophthalmology; (330.2210) Vision - eye movements
7.  Chemical management in fungicide sensivity of Mycosphaerella fijiensis collected from banana fields in México 
Brazilian Journal of Microbiology  2014;45(1):359-364.
The chemical management of the black leaf streak disease in banana caused by Mycosphaerella fijiensis (Morelet) requires numerous applications of fungicides per year. However this has led to fungicide resistance in the field. The present study evaluated the activities of six fungicides against the mycelial growth by determination of EC50 values of strains collected from fields with different fungicide management programs: Rustic management (RM) without applications and Intensive management (IM) more than 25 fungicide application/year. Results showed a decreased sensitivity to all fungicides in isolates collected from IM. Means of EC50 values in mg L−1 for RM and IM were: 13.25 ± 18.24 and 51.58 ± 46.14 for azoxystrobin, 81.40 ± 56.50 and 1.8575 ± 2.11 for carbendazim, 1.225 ± 0.945 and 10.01 ± 8.55 for propiconazole, 220 ± 67.66 vs. 368 ± 62.76 for vinclozolin, 9.862 ± 3.24 and 54.5 ± 21.08 for fludioxonil, 49.2125 ± 34.11 and 112.25 ± 51.20 for mancozeb. A molecular analysis for β-tubulin revealed a mutation at codon 198 in these strains having an EC50 greater than 10 mg L−1 for carbendazim. Our data indicate a consistency between fungicide resistance and intensive chemical management in banana fields, however indicative values for resistance were also found in strains collected from rustic fields, suggesting that proximity among fields may be causing a fungus interchange, where rustic fields are breeding grounds for development of resistant strains. Urgent actions are required in order to avoid fungicide resistance in Mexican populations of M. fijiensis due to fungicide management practices.
PMCID: PMC4059323  PMID: 24948956
banana; fungicides; Mycosphaerella fijiensis; sensitivity; Sigatoka
8.  Cardioprotection by S-nitrosation of a cysteine switch on mitochondrial complex I 
Nature medicine  2013;19(6):753-759.
Oxidative damage from elevated production of reactive oxygen species (ROS) contributes to ischemia-reperfusion injury in myocardial infarction and stroke. The mechanism by which the increase in ROS occurs is not known, and it is unclear how this increase can be prevented. A wide variety of nitric oxide donors and S-nitrosating agents protect the ischemic myocardium from infarction, but the responsible mechanisms are unclear1–6. Here we used a mitochondria-selective S-nitrosating agent, MitoSNO, to determine how mitochondrial S-nitrosation at the reperfusion phase of myocardial infarction is cardioprotective in vivo in mice. We found that protection is due to the S-nitrosation of mitochondrial complex I, which is the entry point for electrons from NADH into the respiratory chain. Reversible S-nitrosation of complex I slows the reactivation of mitochondria during the crucial first minutes of the reperfusion of ischemic tissue, thereby decreasing ROS production, oxidative damage and tissue necrosis. Inhibition of complex I is afforded by the selective S-nitrosation of Cys39 on the ND3 subunit, which becomes susceptible to modification only after ischemia. Our results identify rapid complex I reactivation as a central pathological feature of ischemia-reperfusion injury and show that preventing this reactivation by modification of a cysteine switch is a robust cardioprotective mechanism and hence a rational therapeutic strategy.
doi:10.1038/nm.3212
PMCID: PMC4019998  PMID: 23708290
9.  Clinically-relevant consecutive treatment with isoproterenol and adenosine protects the failing heart against ischaemia and reperfusion 
Background
Consecutive treatment of normal heart with a high dose of isoproterenol and adenosine (Iso/Ade treatment), confers strong protection against ischaemia/reperfusion injury. In preparation for translation of this cardioprotective strategy into clinical practice during heart surgery, we further optimised conditions for this intervention using a clinically-relevant dose of Iso and determined its cardioprotective efficacy in hearts isolated from a model of surgically-induced heart failure.
Methods
Isolated Langendorff-perfused rat hearts were treated sequentially with 5 nM Iso and 30 μM Ade followed by different durations of washout prior to 30 min global ischaemia and 2 hrs reperfusion. Reperfusion injury was assessed by measuring haemodynamic function, lactate dehydrogenase (LDH) release and infarct size. Protein kinase C (PKC) activity and glycogen content were measured in hearts after the treatment. In a separate group of hearts, Cyclosporine A (CsA), a mitochondria permeability transition pore (MPTP) inhibitor, was added with Iso/Ade. Failing hearts extracted after 16 weeks of ligation of left coronary artery in 2 months old rats were also subjected to Iso/Ade treatment followed by ischaemia/reperfusion.
Results
Recovery of the rate pressure product (RPP) in Iso/Ade-treated hearts was significantly higher than in controls. Thus in Iso/Ade treated hearts with 5 nM Iso and no washout period, RPP recovery was 76.3 ± 6.9% of initial value vs. 28.5 ± 5.2% in controls. This was associated with a 3 fold reduction in LDH release irrespective to the duration of the washout period. Hearts with no washout of the drugs (Ade) had least infarct size, highest PKC activity and also showed reduced glycogen content. Cardioprotection with CsA was not additive to the effect of Iso/Ade treatment. Iso/Ade treatment conferred significant protection to failing hearts. Thus, RPP recovery in failing hearts subjected to the treatment was 69.0 ± 16.3% while in Control hearts 19.7 ± 4.0%. LDH release in these hearts was also 3 fold lower compared to Control.
Conclusions
Consecutive Iso/Ade treatment of normal heart can be effective at clinically-relevant doses and this effect appears to be mediated by glycogen depletion and inhibition of MPTP. This intervention protects clinically relevant failing heart model making it a promising candidate for clinical use.
doi:10.1186/1479-5876-12-139
PMCID: PMC4045901  PMID: 24885907
Ischaemia/reperfusion; Cardioprotection; Isoproterenol; Adenosine; Heart failure; Mitochondria permeability transition pore
10.  Low Cardiac Output Secondary to a Malpositioned Umbilical Venous Catheter: Value of Targeted Neonatal Echocardiography 
AJP Reports  2014;4(1):23-28.
Systemic hypotension is common in very low birthweight preterm infants but the nature of the precipitating cause may be unclear. Targeted neonatal echocardiography (TnEcho) is being increasingly used to support hemodynamic decisions in the neonatal intensive care unit (NICU), including identifying impairments in the transitional circulation of preterm infants, providing timely re-evaluation after institution of therapies and evaluating the placement of indwelling catheters. We present a case of a preterm infant with systemic hypotension and low cardiac output secondary to a large transatrial shunt induced by a malpositioned umbilical venous catheter. Repositioning of the line led to resolution of the hemodynamic disturbance and clinical instability, highlighting the utility of TnEcho in the NICU.
doi:10.1055/s-0034-1368090
PMCID: PMC4078164  PMID: 25032055
targeted neonatal echocardiography; umbilical venous catheter; low cardiac output; atrial septal defect
11.  Stimulation of ICa by basal PKA activity is facilitated by caveolin-3 in cardiac ventricular myocytes☆ 
L-type Ca channels (LTCC), which play a key role in cardiac excitation–contraction coupling, are located predominantly at the transverse (t-) tubules in ventricular myocytes. Caveolae and the protein caveolin-3 (Cav-3) are also present at the t-tubules and have been implicated in localizing a number of signaling molecules, including protein kinase A (PKA) and β2-adrenoceptors. The present study investigated whether disruption of Cav-3 binding to its endogenous binding partners influenced LTCC activity. Ventricular myocytes were isolated from male Wistar rats and LTCC current (ICa) recorded using the whole-cell patch-clamp technique. Incubation of myocytes with a membrane-permeable peptide representing the scaffolding domain of Cav-3 (C3SD) reduced basal ICa amplitude in intact, but not detubulated, myocytes, and attenuated the stimulatory effects of the β2-adrenergic agonist zinterol on ICa. The PKA inhibitor H-89 also reduced basal ICa; however, the inhibitory effects of C3SD and H-89 on basal ICa amplitude were not summative. Under control conditions, myocytes stained with antibody against phosphorylated LTCC (pLTCC) displayed a striated pattern, presumably reflecting localization at the t-tubules. Both C3SD and H-89 reduced pLTCC staining at the z-lines but did not affect staining of total LTCC or Cav-3. These data are consistent with the idea that the effects of C3SD and H-89 share a common pathway, which involves PKA and is maximally inhibited by H-89, and suggest that Cav-3 plays an important role in mediating stimulation of ICa at the t-tubules via PKA-induced phosphorylation under basal conditions, and in response to β2-adrenoceptor stimulation.
Graphical abstract
Highlights
•Basal L type calcium current was reduced by interfering with caveolin-3 binding.•L type calcium current is tonically regulated by PKA phosphorylation.•Interfering with caveolin-3 binding reduced beta2 adrenergic stimulation of ICa.
doi:10.1016/j.yjmcc.2013.12.026
PMCID: PMC3980375  PMID: 24412535
t-tubules; Ca; Phosphorylation; β2-Adrenoceptors
12.  A mitochondria-targeted mass spectrometry probe to detect glyoxals: implications for diabetes☆ 
Free Radical Biology & Medicine  2014;67(100):437-450.
The glycation of protein and nucleic acids that occurs as a consequence of hyperglycemia disrupts cell function and contributes to many pathologies, including those associated with diabetes and aging. Intracellular glycation occurs after the generation of the reactive 1,2-dicarbonyls methylglyoxal and glyoxal, and disruption of mitochondrial function is associated with hyperglycemia. However, the contribution of these reactive dicarbonyls to mitochondrial damage in pathology is unclear owing to uncertainties about their levels within mitochondria in cells and in vivo. To address this we have developed a mitochondria-targeted reagent (MitoG) designed to assess the levels of mitochondrial dicarbonyls within cells. MitoG comprises a lipophilic triphenylphosphonium cationic function, which directs the molecules to mitochondria within cells, and an o-phenylenediamine moiety that reacts with dicarbonyls to give distinctive and stable products. The extent of accumulation of these diagnostic heterocyclic products can be readily and sensitively quantified by liquid chromatography–tandem mass spectrometry, enabling changes to be determined. Using the MitoG-based analysis we assessed the formation of methylglyoxal and glyoxal in response to hyperglycemia in cells in culture and in the Akita mouse model of diabetes in vivo. These findings indicated that the levels of methylglyoxal and glyoxal within mitochondria increase during hyperglycemia both in cells and in vivo, suggesting that they can contribute to the pathological mitochondrial dysfunction that occurs in diabetes and aging.
Highlights
•A mitochondria-targeted mass spectrometric probe, MitoG, has been developed to measure glyoxal and methylglyoxal.•Using MitoG we show that mitochondrial glyoxal and methylglyoxal can be measured in hyperglycemic cells.•MitoG can also be used in vivo to infer mitochondrial glyoxal and methylglyoxal production in a mouse model of type I diabetes.•These findings suggest that the accumulation of glyoxal and methylglyoxal within mitochondria may contribute to mitochondrial dysfunction in diabetes.
doi:10.1016/j.freeradbiomed.2013.11.025
PMCID: PMC3978666  PMID: 24316194
Mitochondria; Exomarker; Methylglyoxal; Glyoxal; Hyperglycemia; MitoG; Free radicals
13.  Plant immunity. Methods and protocols 
Annals of Botany  2013;111(1):viii.
doi:10.1093/aob/mcs272
PMCID: PMC3523659
14.  Balloon Dilators for Labor Induction: a Historical Review 
A number of recent articles attribute the origin of the use of cervical balloon dilation in the induction of labor to either Barnes in the 1860s or Embrey and Mollison in the 1960s. This review examines the historical record and reveals that, based on current practice attribution should rather be made to two contemporaries of Barnes: the Storer and Mattei. More importantly, Storer’s warning about the rubber used in dilators was ignored, leading to decades of possibly unnecessary deaths following childbirth. To conduct this study key search terms for PubMed, Google Scholar and the website of the University of Ryerson were utilized as “Barnes”, “Woodman”, “balloon dilation”, “balloon catheter”, “foley”, “colpeurynter”, “cervix uteri” and “induction.” Subsequent analysis was done on downloaded articles using BibDesk.
PMCID: PMC3885146  PMID: 24427487
balloon dilation; cervical dilation; foley catheter; hydrostatic dilator; induction of labor
15.  Glycolytic ATP Fuels the Plasma Membrane Calcium Pump Critical for Pancreatic Cancer Cell Survival* 
The Journal of Biological Chemistry  2013;288(50):36007-36019.
Background: Pancreatic cancer cells exhibit up-regulated glycolysis.
Results: Inhibition of glycolysis, but not mitochondrial metabolism, induced ATP depletion, plasma membrane calcium pump (PMCA) inhibition, Ca2+ overload, and cell death.
Conclusion: Glycolytic ATP fuels the PMCA in pancreatic cancer.
Significance: Glycolytic regulation of the PMCA may represent a novel chemotherapeutic target for pancreatic cancer.
Pancreatic cancer is an aggressive cancer with poor prognosis and limited treatment options. Cancer cells rapidly proliferate and are resistant to cell death due, in part, to a shift from mitochondrial metabolism to glycolysis. We hypothesized that this shift is important in regulating cytosolic Ca2+ ([Ca2+]i), as the ATP-dependent plasma membrane Ca2+ ATPase (PMCA) is critical for maintaining low [Ca2+]i and thus cell survival. The present study aimed to determine the relative contribution of mitochondrial versus glycolytic ATP in fuelling the PMCA in human pancreatic cancer cells. We report that glycolytic inhibition induced profound ATP depletion, PMCA inhibition, [Ca2+]i overload, and cell death in PANC1 and MIA PaCa-2 cells. Conversely, inhibition of mitochondrial metabolism had no effect, suggesting that glycolytic ATP is critical for [Ca2+]i homeostasis and thus survival. Targeting the glycolytic regulation of the PMCA may, therefore, be an effective strategy for selectively killing pancreatic cancer while sparing healthy cells.
doi:10.1074/jbc.M113.502948
PMCID: PMC3861649  PMID: 24158437
ATP; Calcium ATPase; Calcium Signaling; Cell Death; Glycolysis; Metabolism; Pancreatic Cancer; PMCA; Warburg; Calcium Overload
16.  A simplified cervix model in response to induction balloon in pre-labour 
Background
Induction of labour is poorly understood even though it is performed in 20% of births in the United States. One method of induction, the balloon dilator applied with traction to the interior os of the cervix, engages a softening process, permitting dilation and effacement to proceed until the beginning of active labour. The purpose of this work is to develop a simple model capable of reproducing the dilation and effacement effect in the presence of a balloon.
Methods
The cervix, anchored by the uterus and the endopelvic fascia was modelled in pre-labour. The spring-loaded, double sliding-joint, double pin-joint mechanism model was developed with a Modelica-compatible system, MapleSoft MapleSim 6.1, with a stiff Rosenbrock solver and 1E-4 absolute and relative tolerances. Total simulation time for pre-labour was seven hours and simulations ended at 4.50 cm dilation diameter and 2.25 cm effacement.
Results
Three spring configurations were tested: one pin joint, one sliding joint and combined pin-joint-sliding-joint. Feedback, based on dilation speed modulated the spring values, permitting controlled dilation. Dilation diameter speed was maintained at 0.692 cm·hr-1 over the majority of the simulation time. In the sliding-joint-only mode the maximum spring constant value was 23800 N·m-1. In pin-joint-only the maximum spring constant value was 0.41 N·m·rad-1. With a sliding-joint-pin-joint pair the maximum spring constants are 2000 N·m-1 and 0.41 N·m·rad-1, respectively.
Conclusions
The model, a simplified one-quarter version of the cervix, is capable of maintaining near-constant dilation rates, similar to published clinical observations for pre-labour. Lowest spring constant values are achieved when two springs are used, but nearly identical tracking of dilation speed can be achieved with only a pin joint spring. Initial and final values for effacement and dilation also match published clinical observations. These results provide a framework for development of electro-mechanical phantoms for induction training, as well as dilator testing and development.
doi:10.1186/1742-4682-10-58
PMCID: PMC3850663  PMID: 24070547
Balloon dilator; Cervix; Pre-labour; Latent phase of labour; Labour induction; Dilation; Effacement
17.  Screw Placement Accuracy for Minimally Invasive Transforaminal Lumbar Interbody Fusion Surgery: A Study on 3-D Neuronavigation-Guided Surgery 
Global Spine Journal  2012;2(3):143-152.
Purpose The aim of this study was to assess the impact of 3-D navigation for pedicle screw placement accuracy in minimally invasive transverse lumbar interbody fusion (MIS-TLIF).
Methods A retrospective review of 52 patients who had MIS-TLIF assisted with 3D navigation is presented. Clinical outcomes were assessed with the Oswestry Disability Index (ODI), Visual Analog Scales (VAS), and MacNab scores. Radiographic outcomes were assessed using X-rays and thin-slice computed tomography.
Result The mean age was 56.5 years, and 172 screws were implanted with 16 pedicle breaches (91.0% accuracy rate). Radiographic fusion rate at a mean follow-up of 15.6 months was 87.23%. No revision surgeries were required. The mean improvement in the VAS back pain, VAS leg pain, and ODI at 11.3 months follow-up was 4.3, 4.5, and 26.8 points, respectively. At last follow-up the mean postoperative disc height gain was 4.92 mm and the mean postoperative disc angle gain was 2.79 degrees. At L5–S1 level, there was a significant correlation between a greater disc space height gain and a lower VAS leg score.
Conclusion Our data support that application of 3-D navigation in MIS-TLIF is associated with a high level of accuracy in the pedicle screw placement.
doi:10.1055/s-0032-1326949
PMCID: PMC3864504  PMID: 24353961
minimally invasive spine surgery; transforaminal lumbar interbody fusion; 3D-NAV; neuronavigation; pedicle screw
18.  Nickel inhibits β-1 adrenoceptor mediated activation of cardiac CFTR chloride channels 
Highlights
► We report the block of the β-adrenoceptor-activated cardiac CFTR Cl− current by Ni2+. ► Extracellular Ni2+ inhibits the current activated by β1-adrenoceptors in a concentration-dependent manner. ► The action of Ni2+ is insensitive to β2-blockade. ► Ni2+ does not affect the β-adrenoceptor-activated current from the intracellular side. ► The data are consistent with an action of Ni2+ at the β1-adrenoceptor from the external side.
Cardiac ventricular myocytes exhibit a protein kinase A-dependent Cl− current (ICl.PKA) mediated by the cystic fibrosis transmembrane conductance regulator (CFTR). There is conflicting evidence regarding the ability of the divalent cation nickel (Ni2+), which has been used widely in vitro in the study of other cardiac ionic conductances, to inhibit ICl.PKA. Here the action of Ni2+ on ICl.PKA activated by β-adrenergic stimulation has been elucidated. Whole-cell patch-clamp recordings were made from rabbit isolated ventricular myocytes. Externally applied Ni2+ blocked ICl.PKA activated by 1 μM isoprenaline with a log IC50 (M) of −4.107 ± 0.075 (IC50 = 78.1 μM) at +100 mV and −4.322 ± 0.107 (IC50 = 47.6 μM) at −100 mV. Thus, the block of ICl.PKA by Ni2+ was not strongly voltage dependent. Ni2+ applied internally via the patch-pipette was ineffective at inhibiting isoprenaline-activated ICl,PKA, but in the same experiments the current was suppressed by external Ni2+ application, indicative of an external site of Ni2+ action. In the presence of 1 μM atenolol isoprenaline was ineffective at activating ICl.PKA, but in the presence of the β2-adrenoceptor inhibitor ICI 118,551 isoprenaline still activated Ni2+-sensitive ICl.PKA. Collectively, these data demonstrate that Ni2+ ions produce marked inhibition of β1-adrenoceptor activated ventricular ICl.PKA at submillimolar [Ni2+]: an action that is likely to involve an interaction between Ni2+ and β1-adrenoceptors. The concentration-dependence for ICl.PKA inhibition seen here indicates the potential for confounding effects on ICl,PKA to occur even at comparatively low Ni2+ concentrations, when Ni2+ is used to study other cardiac ionic currents under conditions of β-adrenergic agonism.
doi:10.1016/j.bbrc.2013.01.087
PMCID: PMC3686155  PMID: 23376720
Rabbit cardiomyocytes; PKA-dependent Cl− current; CFTR; CFTR-inhibitor; Nickel; Ni2+
19.  Molecular Basis of Calcium-Sensitizing and Desensitizing Mutations of the Human Cardiac Troponin C Regulatory Domain: A Multi-Scale Simulation Study 
PLoS Computational Biology  2012;8(11):e1002777.
Troponin C (TnC) is implicated in the initiation of myocyte contraction via binding of cytosolic and subsequent recognition of the Troponin I switch peptide. Mutations of the cardiac TnC N-terminal regulatory domain have been shown to alter both calcium binding and myofilament force generation. We have performed molecular dynamics simulations of engineered TnC variants that increase or decrease sensitivity, in order to understand the structural basis of their impact on TnC function. We will use the distinction for mutants that are associated with increased affinity and for those mutants with reduced affinity. Our studies demonstrate that for GOF mutants V44Q and L48Q, the structure of the physiologically-active site II binding site in the -free (apo) state closely resembled the -bound (holo) state. In contrast, site II is very labile for LOF mutants E40A and V79Q in the apo form and bears little resemblance with the holo conformation. We hypothesize that these phenomena contribute to the increased association rate, , for the GOF mutants relative to LOF. Furthermore, we observe significant positive and negative positional correlations between helices in the GOF holo mutants that are not found in the LOF mutants. We anticipate these correlations may contribute either directly to affinity or indirectly through TnI association. Our observations based on the structure and dynamics of mutant TnC provide rationale for binding trends observed in GOF and LOF mutants and will guide the development of inotropic drugs that target TnC.
Author Summary
Muscle cells contract using a network of thread-like protein assemblies called myofilaments. Contraction is preceded by a signal that causes calcium to rush into the cell cytosol, where it can freely diffuse to and bind the myofilament proteins. Troponin C, a calcium sensor located on the thin filament, initiates and regulates the cascade of changes resulting in the generation of force by the thin and thick filaments comprising the myofilament lattice. In heart tissue, pathological conditions known as dilated and hypertrophic cardiomyopathies (DCM and HCM, respectively) are in part associated with abnormalities in the ability of the myofilaments to generate force at normal calcium concentrations. Manipulation of Troponin C calcium-binding through protein engineering and pharmaceutical intervention has thus attracted considerable attention as a therapeutic strategy for ameliorating these cardiac defects. In this study, we uncover a molecular basis of altered calcium handling for several engineered Troponin C variants, which provides further insight into tuning its control of myofilament contraction.
doi:10.1371/journal.pcbi.1002777
PMCID: PMC3510055  PMID: 23209387
20.  Acute desensitization of acetylcholine and endothelin-1 activated inward rectifier K+ current in myocytes from the cardiac atrioventricular node 
Highlights
► ACh and ET-1 activate a K+ current in cardiac atrioventricular nodal cells. ► Tertiapin-Q sensitive IKACh activated via M2 receptors shows bi-exponential ‘fade’. ► ET-1 activates a similar current that also fades. ► The fade reflects desensitization rather than altered K+ ion driving force. ► Acetylcholine is able to cross-desensitize the AVN cell response to endothelin-1.
The atrioventricular node (AVN) is a vital component of the pacemaker-conduction system of the heart, co-ordinating conduction of electrical excitation from cardiac atria to ventricles and acting as a secondary pacemaker. The electrical behaviour of the AVN is modulated by vagal activity via activation of muscarinic potassium current, IKACh. However, it is not yet known if this response exhibits ‘fade’ or desensitization in the AVN, as established for the heart’s primary pacemaker – the sinoatrial node. In this study, acute activation of IKACh in rabbit single AVN cells was investigated using whole-cell patch clamp at 37 °C. 0.1–1 μM acetylcholine (ACh) rapidly activated a robust IKACh in AVN myocytes during a descending voltage-ramp protocol. This response was inhibited by tertiapin-Q (TQ; 300 nM) and by the M2 muscarinic ACh receptor antagonist AFDX-116 (1 μM). During sustained ACh exposure the elicited IKACh exhibited bi-exponential fade (τf of 2.0 s and τs 76.9 s at −120 mV; 1 μM ACh). 10 nM ET-1 elicited a current similar to IKACh, which faded with a mono-exponential time-course (τ of 52.6 s at −120 mV). When ET-1 was applied following ACh, the ET-1 activated response was greatly attenuated, demonstrating that ACh could desensitize the response to ET-1. For neither ACh nor ET-1 was the rate of current fade dependent upon the initial response magnitude, which is inconsistent with K+ flux mediated changes in electrochemical driving force as the underlying mechanism. Collectively, these findings demonstrate that TQ sensitive inwardly rectifying K+ current in cardiac AVN cells, elicited by M2 muscarinic receptor or ET-1 receptor activation, exhibits fade due to rapid desensitization.
doi:10.1016/j.bbrc.2012.05.148
PMCID: PMC3400056  PMID: 22683635
Acetylcholine (ACh); Atrioventricular node; AV node; AVN; Endothelin-1 (ET-1); GIRK; IKACh; Inward rectifier; Muscarinic potassium current; Tertiapin-Q
21.  Tissue-engineered total disc replacement: final outcomes of a murine caudal disc in vivo study  
Selected abstracts delivered at the 9th Annual AOSpine North America Fellows Forum
Consistent with EBSJ's commitment to fostering quality research, we are pleased to feature some of the most highly rated abstracts from the 9th Annual AOSpine North America Fellows Forum in Banff, Canada. Enhancing the quality of evidence in spine care means acknowledging and supporting the efforts of young researchers within our AOSpine North America network. We look forward to seeing more from these promising researchers in the future.
doi:10.1055/s-0031-1274758
PMCID: PMC3506141  PMID: 23230409
22.  Modulation by Endothelin-1 of Spontaneous Activity and Membrane Currents of Atrioventricular Node Myocytes from the Rabbit Heart 
PLoS ONE  2012;7(3):e33448.
Background
The atrioventricular node (AVN) is a key component of the cardiac pacemaker-conduction system. Although it is known that receptors for the peptide hormone endothelin-1 (ET-1) are expressed in the AVN, there is very little information available on the modulatory effects of ET-1 on AVN electrophysiology. This study characterises for the first time acute modulatory effects of ET-1 on AVN cellular electrophysiology.
Methods
Electrophysiological experiments were conducted in which recordings were made from rabbit isolated AVN cells at 35–37°C using the whole-cell patch clamp recording technique.
Results
Application of ET-1 (10 nM) to spontaneously active AVN cells led rapidly (within ∼13 s) to membrane potential hyperpolarisation and cessation of spontaneous action potentials (APs). This effect was prevented by pre-application of the ETA receptor inhibitor BQ-123 (1 µM) and was not mimicked by the ETB receptor agonist IRL-1620 (300 nM). In whole-cell voltage-clamp experiments, ET-1 partially inhibited L-type calcium current (ICa,L) and rapid delayed rectifier K+ current (IKr), whilst it transiently activated the hyperpolarisation-activated current (If) at voltages negative to the pacemaking range, and activated an inwardly rectifying current that was inhibited by both tertiapin-Q (300 nM) and Ba2+ ions (2 mM); each of these effects was sensitive to ETA receptor inhibition. In cells exposed to tertiapin-Q, ET-1 application did not produce membrane potential hyperpolarisation or immediate cessation of spontaneous activity; instead, there was a progressive decline in AP amplitude and depolarisation of maximum diastolic potential.
Conclusions
Acutely applied ET-1 exerts a direct modulatory effect on AVN cell electrophysiology. The dominant effect of ET-1 in this study was activation of a tertiapin-Q sensitive inwardly rectifying K+ current via ETA receptors, which led rapidly to cell quiescence.
doi:10.1371/journal.pone.0033448
PMCID: PMC3315568  PMID: 22479400
23.  Selective Uncoupling of Individual Mitochondria within a Cell Using a Mitochondria-Targeted Photoactivated Protonophore 
Depolarization of an individual mitochondrion or small clusters of mitochondria within cells has been achieved using a photoactivatable probe. The probe is targeted to the matrix of the mitochondrion by an alkyltriphenylphosphonium lipophilic cation and releases the protonophore 2,4-dinitrophenol locally in predetermined regions in response to directed irradiation with UV light via a local photolysis system. This also provides a proof of principle for the general temporally and spatially controlled release of bioactive molecules, pharmacophores, or toxins to mitochondria with tissue, cell, or mitochondrion specificity.
doi:10.1021/ja2077922
PMCID: PMC3260739  PMID: 22239373
24.  ADAPTIVE FINITE ELEMENT MODELING TECHNIQUES FOR THE POISSON-BOLTZMANN EQUATION 
We consider the design of an effective and reliable adaptive finite element method (AFEM) for the nonlinear Poisson-Boltzmann equation (PBE). We first examine the two-term regularization technique for the continuous problem recently proposed by Chen, Holst, and Xu based on the removal of the singular electrostatic potential inside biomolecules; this technique made possible the development of the first complete solution and approximation theory for the Poisson-Boltzmann equation, the first provably convergent discretization, and also allowed for the development of a provably convergent AFEM. However, in practical implementation, this two-term regularization exhibits numerical instability. Therefore, we examine a variation of this regularization technique which can be shown to be less susceptible to such instability. We establish a priori estimates and other basic results for the continuous regularized problem, as well as for Galerkin finite element approximations. We show that the new approach produces regularized continuous and discrete problems with the same mathematical advantages of the original regularization. We then design an AFEM scheme for the new regularized problem, and show that the resulting AFEM scheme is accurate and reliable, by proving a contraction result for the error. This result, which is one of the first results of this type for nonlinear elliptic problems, is based on using continuous and discrete a priori L∞ estimates to establish quasi-orthogonality. To provide a high-quality geometric model as input to the AFEM algorithm, we also describe a class of feature-preserving adaptive mesh generation algorithms designed specifically for constructing meshes of biomolecular structures, based on the intrinsic local structure tensor of the molecular surface. All of the algorithms described in the article are implemented in the Finite Element Toolkit (FETK), developed and maintained at UCSD. The stability advantages of the new regularization scheme are demonstrated with FETK through comparisons with the original regularization approach for a model problem. The convergence and accuracy of the overall AFEM algorithm is also illustrated by numerical approximation of electrostatic solvation energy for an insulin protein.
doi:10.4208/cicp.081009.130611a
PMCID: PMC3178275  PMID: 21949541
Poisson-Boltzmann equation; semi-linear partial differential equations; super-critical nonlinearity; singularity; a priori L∞ estimates; existence; uniqueness; well-posedness; Galerkin methods; discrete a priori L∞ estimates; quasi-optimal a priori error estimates; adaptive finite methods; contraction; convergence; optimality; surface and volume mesh generation; mesh improvement and decimation
25.  Biological intervertebral disc replacement: an in vivo model and comparison of two surgical techniques to approach the rat caudal disc 
Study design: Prospective randomized animal study.
Objective: To determine a surgical technique for reproducible and functional intervertebral disc replacement in an orthotopic animal model.
Methods: The caudal 3/4 intervertebral disc (IVD) of the rat tail was approached by two surgical techniques: blunt dissection, stripping and retracting (Technique 1) or incising and repairing (Technique 2) the dorsal longitudinal tendons. The intervertebral disc was dissected and removed, and then either discarded or reinserted. Outcome measures were perioperative complications, spontaneous tail movement, 7T MRI (T1- and T2-sequences for measurement of disc space height (DSH) and disc hydration). Microcomputed tomographic imaging (micro CT) was additionally performed postmortem.
Results: No vascular injuries occurred and no systemic or local infections were observed over the course of 1 month. Tail movements were maintained. With tendon retraction (Technique 1) gross loss of DSH occurred with both discectomy and reinsertion. Tendon division (Technique 2) maintained DSH with IVD reinsertion but not without. The DSH was demonstrated on MRI measurement. A new scoring system to assess IVD appearances was described.
Conclusions: The rat tail model, with a tendon dividing surgical technique, can function as an orthotopic animal model for IVD research. Mechanical stimulation is maintained by preserved tail movements. 7T MRI is a feasible modality for longitudinal monitoring for the rat caudal disc.
doi:10.1055/s-0030-1267084
PMCID: PMC3427968  PMID: 22956934

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